Cytokine production inhibitors, agents for protecting and promoting liver funtion, anti-inflammatory agents, immunosuppressants, drugs, cosmetics, foods and food materials

ABSTRACT

Highly safe cytokine production inhibitors, agents for protecting and promoting liver function, anti-inflammatory agents and immunosuppressants. Brazilian licorice extract and periandrins, which are constituents thereof, have excellent characteristics of showing effects of inhibiting cytokine production and protecting and promoting liver function, an anti-inflammatory effect and an immunosuppressive effect without any harmful side effects. Thus use of Brazilian licorice extract or periandrins as cytoline production inhibitors makes it possible to inhibit inflammations in various diseases such as rheumatoid arthritis. These substances are also usable as agents for protecting and promoting liver function, anti-inflammatory agents and immunosuppressants. Moreover, foods, cosmetics, sweeteners and food materials containing Brazilian licorice extract exert the effects as described above.

TECHNICAL FIELD OF THE INVENTION

[0001] This invention relates to cytokine production inhibitors, agentsfor protecting and promoting liver function, anti-inflammatory agents,immunosuppressants, drugs, cosmetics, foods, and food materials.

BACKGROUND OF THE INVENTION

[0002] Inflammation by autoimmune diseases like rheumatism, allergiesand atopies is promoted by production of migration enhancement factorsof leukocytes called cytokine.

[0003] In order to inhibit such cytokine production, steroids such asprednisolone have been conventionally used.

[0004] However, steroids have strong side effects. Therefore, repeatedadministration of the same is difficult.

[0005] To make matters worse, there is no report on any component ofedible plants and galenicals which shows strong effect of inhibitingcytokine production even at the basic research level.

[0006] In short, hardly any medical agent has been existed which caninhibit cytokine production without any side effects, and such an agenthas been expected to be discovered.

[0007] Licorice (leguminous perennial herb) and glycyrrhizin which is acomponent thereof have been used for an anti-inflammatory agent,anti-allergic agent and agent for protecting liver function, whichpromote detoxication and tissue recovery.

[0008] However, since glycyrrhizin inhibits corticosteroid metabolism,pseudoaldosteronism (edema with sodium retention, hypertension) may bedeveloped as a side effect.

[0009] Therefore, a licorice cannot be administered over a long periodto a patient having kidney diseases or hypertension.

[0010] Accordingly, a medical agent which is as effective as licoricebut which does not develop any side effect has been expected.

SUMMARY OF THE INVENTION

[0011] One object of the present invention is to provide a medical agentwhich exerts effects of inhibiting cytokine production, protecting andpromoting liver function, an anti-inflammatory effect, and animmunosuppressive effect, and which hardly causes any side effect.Another object of the present invention is to provide drugs, cosmetics,foods, and food materials to which the aforementioned medical agent isapplied.

[0012] Heretofore, periandrins and periandradulcins have been reportedas main components of a Brazilian licorice. These are both glycosidesstructurally similar to glycyrrhizin which is a component of a licorice,and are expected to act similar to glycyrrhizin. However, hardly anyreport has been made in relation to their physiological activities.

[0013] The inventors discovered that Brazilian licorice extract andperiandrins, which are constituents thereof, have an anti-inflammatoryeffect and an immunosuppressive effect, and accomplished the presentinvention.

[0014] Brazilian licorice extract is, for example, obtained usingextraction vehicles such as ethyl alcohol.

[0015] (1) The invention set forth in claim 1 provides a cytokineproduction inhibitor comprising periandrins as an active ingredient.

[0016] Since the periandrins show an effect of inhibiting cytokineproduction, the cytokine production inhibitor of the present inventioninhibits cytokine production and acts as an anti-inflammatory agent.

[0017] Accordingly, the cytokine production inhibitor of the presentinvention can inhibit inflammation developed by cell disorders (forexample, by bacteria, toxic elements in medicine and alcohol),autoimmune diseases (such as rheumatism (rheumatoid arthritis, forexample), systemic lupus erythematosus, collagenosis, etc.), atopies,allergies, and pollinosis.

[0018] Furthermore, periandrins do not inhibit activation of 11β-hydroxysteroid dehydrogenase.

[0019] If activation of 11 β-hydroxysteroid dehydrogenase is inhibited,corticosteroid metabolism is also inhibited, resulting in thatpseudoaldosteronism (edema with sodium retention, hypertension) mayoccur.

[0020] Accordingly, the cytokine production inhibitor of the presentinvention comprising periandrins does not cause pseudoaldosteronism andcan be administered, for example, even to a patient having kidneydiseases and hypertension over a long period.

[0021] The cytokine production inhibitor of the present invention can beproduced by compounding periandrins obtained by separating a componentheld by adsorption chromatography carrier from the Brazilian licoriceextract, for example.

[0022] In this case, since polysaccharide contained in the Brazilianlicorice extract is not entered into the cytokine production inhibitor,it is difficult for the cytokine production inhibitor to get moldy.

[0023] Examples of the aforementioned periandrins are periandrin I(PD-I), periandrin II (PD-II), periandrin III (PD-III), periandrin IV(PD-IV), etc.

[0024] Cytokine is a cellular glycoprotein, and an endogenous substancein vivo which acts on the other cells. It is a general term forinterleukin, interferon, lymphokine, monokine, and tumor necrosisfactors.

[0025] When cells are damaged by bacteria, toxic elements in medicine,alcohol, etc., or in case of autoimmune diseases such as rheumatism,collagenosis, etc., phagocyte such as macrophage, etc. is activated andautopepsia occurs, resulting in that inflammation is developed. At thesame time, cytokine which is a type of migration enhancement factors ofleukocytes is produced. However, cytokine has a number of factors whichaccelerates immunoreaction. Therefore, if cytokine is produced in alarge amount, it accelerates aggregation of phagocyte and furtherworsens the inflammation.

[0026] (2) The invention set forth in claim 2 provides a cytokineproduction inhibitor containing one of Brazilian licorice extract andone or more constituents contained in the extract as an activeingredient.

[0027] The cytokine production inhibitor of the present inventioncontains an extract, or one or more constituents contained in the same,obtained by filtering and drying the liquid extracted from a driedrootstalk of a Brazilian licorice with alcohol.

[0028] Brazilian licorice extract inhibits cytokine production, and thusthe cytokine production inhibitor of the present invention inhibitscytokine production and serves as an anti-inflammatory agent.

[0029] Inflammation which is a symptom of diseases such as celldisorders (for example, by bacteria, toxic elements in medicine andalcohol), autoimmune diseases (such as rheumatism (rheumatoid arthritis,for example), systemic lupus erythematosus, collagenosis, etc.),atopies, allergies, and pollinosis is in many cases accelerated bycytokine. Consequently, the cytokine production inhibitor of the presentinvention inhibits inflammation by the aforementioned diseases.

[0030] Brazilian licorice extract, in addition, is on the list of foodadditives in Japan as a sweetener, and is superior in that it developsno harmful side effect.

[0031] (3) The invention set forth in claim 3 provides the cytokineproduction inhibitor claimed in claim 2, one or more constituents of theaforementioned comprise periandrins.

[0032] The cytokine production inhibitor of the present inventioninhibits cytokine production due to periandrins contained in Brazilianlicorice extract.

[0033] Accordingly, the cytokine production inhibitor of the presentinvention, as well as the cytokine production inhibitor claimed in claim1, inhibits inflammation developed by cell disorders, autoimmunediseases, atopies, allergies, and pollinosis, which may be worsened bycytokine.

[0034] This cytokine production inhibitor does not inhibit activation of11 β-hydroxysteroid dehydrogenase, and thus does not causepseudoaldosteronism and can be administered, for example, to a patienthaving kidney diseases and hypertension, over a long period.

[0035] Furthermore, this cytokine production inhibitor has no harmfulside effect. Therefore, it can be administered over a long period intreatment of autoimmune diseases and after organ transplant, forexample, without developing any side effect unlike the conventionalsteroids.

[0036] (4) The invention set forth in claim 4 provides drugs comprisingthe cytokine production inhibitor claimed in one of claims 1-3, as ananti-inflammatory, anti-allergic, or anti-atopic active ingredient.

[0037] The drugs of the present invention comprise periandrins,Brazilian licorice extract, or one or more constituents of the extract.Therefore, it inhibits cytokine production and eases symptoms ofinflammation, allergy, atopy, and pollinosis.

[0038] The drugs of the present invention are also advantageous in thatthey have no side effect.

[0039] The drugs of the present invention can be taken as oral medicine,external preparations, injectable solutions, vaparole, nose drops, eyedrops, etc., for example.

[0040] The drugs of the present invention can be in the form of pill ortablet, liquid, injectable solution, ointment, cream, lotion, aerosol,suppository, etc., for example.

[0041] The drugs of the present invention can compound other ingredientswhich show an anti-inflammatory effect, an anti-allergic effect, or ananti-atopic effect.

[0042] (5) The invention set forth in claim 5 provides cosmeticscomprising the cytokine production inhibitor claimed in one of claims1-3, as an anti-inflammatory, anti-allergic, or anti-atopic activeingredient.

[0043] The cosmetics of the present invention comprise periandrins,Brazilian licorice extract, or one or more constituents of the extract.Therefore, it inhibits cytokine production and eases symptoms ofinflammation, allergy, atopy, and pollinosis.

[0044] Accordingly, use of the cosmetics of the present invention canimprove itches and pains on the skin by allergic symptoms andinflammation. The cosmetics of the present invention can be used even byan atopic person.

[0045] The cosmetics of the present invention are also advantageous inthat they have no side effect.

[0046] The cosmetics of the present invention can be in the form ofcream, ointment, lotion, milky lotion, solid body, powder, etc., forexample.

[0047] Examples of the cosmetics of the present invention are basic skincare products such as lotion, milky lotion, essence, moisturizing cream,etc., sun protection products such as sunscreen cream, sunscreen lotion,sunscreen oil, carmine lotion, etc., makeups such as foundation, eyeliner, mascara, eye shadow, blush-on for cheeks, lipstick, etc.,cleansers such as facial wash, body shampoo, hair shampoo, etc,perfumes, antiperspirant deodorants, etc.

[0048] (6) The invention set forth in claim 6 provides foods comprisingthe cytokine production inhibitor claimed in one of claims 1-3, as ananti-inflammatory, anti-allergic, or anti-atopic active ingredient.

[0049] The foods of the present invention contain periandrins, Brazilianlicorice extract, or one or more constituents of the extract. Therefore,it inhibits cytokine production and eases symptoms of inflammation,allergy, atopy, and pollinosis.

[0050] Accordingly, an intake of the foods of the present invention canimprove various inflammatory symptoms by atopy, pollinosis, etc., forexample.

[0051] The foods of the present invention are also advantageous in thatthey have no side effect.

[0052] The foods of the present invention can be made into oralcompositions such as tea, soft drinks, gums, candies, etc., pastes ofmarine products such as steamed fish pastes, fish sausages, etc., stockfarm products such as sausages, hams, etc., Western-style confectionery,Japanese-style confectionery, noodles such as raw noodles, noodles forboiling, etc., seasonings such as sauces, soy sauces, dips, etc., andgeneral food products such as pickles, prepared foods, etc.

[0053] The foods of the present invention can, as far as the effect ofthe present invention is not deteriorated, compound various ingredientsgenerally used in foods, such as sugar, condensed milk, wheat flour,shortening, salt, glucose sugar, chicken eggs, butter, margarine, starchsyrup, calcium, iron, vitamins, seasonings, spices, etc.

[0054] (7) The invention set forth in claim 7 provides food materialscomprising the cytokine production inhibitor claimed in one of claims1-3, as an anti-inflammatory, anti-allergic, or anti-atopic activeingredient.

[0055] The food materials of the present invention comprise periandrins,Brazilian licorice extract, or one or more constituents of the extract.Therefore, it inhibits cytokine production and eases symptoms ofinflammation, allergy, atopy, and pollinosis.

[0056] Accordingly, an intake of foods compounding the food materials ofthe present invention can improve various inflammatory symptoms byatopy, pollinosis, etc., for example.

[0057] The food materials of the present invention are also advantageousin that they have no side effect.

[0058] The food materials of the present invention can be a sweetener,for example.

[0059] The food materials of the present invention can compound variousingredients generally used in foods (like sugar, condensed milk, wheatflour, shortening, salt, glucose sugar, chicken eggs, butter, margarine,starch syrup, calcium, iron, vitamins, seasonings, spices, etc.).

[0060] The food materials of the present invention can be added tovarious foods and drinks (like oral compositions such as tea, softdrinks, gums, candies, etch, pastes of marine products such as steamedfish pastes, fish sausages, etc., stock farm products such as sausages,hams, etc., Western-style confectionery, Japanese-style confectionery,noodles such as raw noodles, noodles for boiling, etc., seasonings suchas sauces, soy sauces, dips, etc., and general food products such aspickles, prepared foods, etc.).

[0061] (8) The invention set forth in claim 8 provides an agent forprotecting and promoting liver function comprising periandrins as anactive ingredient.

[0062] The periandrins show an effect of protecting and promoting liverfunction, and thus the agent for protecting and promoting liverfunction, comprising periandrins as an active ingredient, of the presentinvention has also an effect of protecting and promoting liver function.

[0063] Cytokine is produced in case that there are any liver disorders.However, the agent for protecting and promoting liver function of thepresent invention inhibits cytokine production by protecting andpromoting liver function and eases symptoms of inflammation, allergy,atopy, and pollinosis, for example.

[0064] Accordingly, the agent for protecting and promoting liverfunction of the present invention is as effective as the conventionallyused licorice extract, and more advantageously, it has no side effectunlike licorice.

[0065] Therefore, the agent for protecting and promoting liver functionof the present invention can be administered over a long period, forexample, to a patient who has kidney diseases and hypertension and towhom long-term administration of licorice is not possible.

[0066] The agent for protecting and promoting liver function of thepresent invention can compound periandrins separated and purified fromBrazilian licorice extract, for example. In this case, sincepolysaccharide contained in Brazilian licorice extract is not enteredinto the agent for protecting and promoting liver function, it isdifficult for the agent for protecting and promoting liver function toget moldy.

[0067] Examples of the aforementioned periandrins are periandrin I(PD-I), periandrin II (PD-II), periandrin III (PD-III), periandrin IV(PD-IV), etc.

[0068] (9) The invention set forth in claim 9 provides an agent forprotecting and promoting liver function comprising one of Brazilianlicorice extract and one or more constituents contained in the extractas an active ingredient.

[0069] The agent for protecting and promoting liver function of thepresent invention comprises extract, or one or more constituentscontained in the same, obtained by filtering and drying liquid extractedfrom a dried rootstalk of a Brazilian licorice with alcohol.

[0070] Brazilian licorice extract shows an effect of protecting andpromoting liver function, and thus the agent for protecting andpromoting liver function, comprising Brazilian licorice extract as anactive ingredient, of the present invention is also effective inprotecting and promoting of liver function.

[0071] Cytokine is produced in case that there are any liver disorders.However, the agent for protecting and promoting liver function of thepresent invention inhibits cytokine production by protecting andpromoting liver function and eases symptoms of inflammation, allergy,atopy, and pollinosis, for example.

[0072] Accordingly, the agent for protecting and promoting liverfunction of the present invention is as effective as the conventionallyused licorice extract. More advantageously, it has no side effect unlikelicorice.

[0073] Therefore, the agent for protecting and promoting liver functionof the present invention can be administered over a long period, forexample, to a patient who has kidney diseases and hypertension and towhom long-term administration of licorice is not possible.

[0074] (1) The invention set forth in claim 10 provides the agent forprotecting and promoting liver function claimed in claim 9, one or moreconstituents of the aforementioned comprise periandrins.

[0075] The agent for protecting and promoting liver function of thepresent invention comprises periandrins contained in the extract as anactive ingredient.

[0076] Therefore, this agent for protecting and promoting liverfunction, as well as the agent for protecting and promoting liverfunction claimed in claim 8, inhibits cytokine production by protectingand promoting liver function and eases symptoms of inflammation,allergy, atopy, and pollinosis, for example.

[0077] Also this agent for protecting and promoting liver function hasno side effect (such as pseudoaldosteronism). Therefore, it can beadministered over a long period, for example, to a patient who haskidney diseases and hypertension and to whom long-term administration oflicorice is not possible.

[0078] (11) The invention set forth in claim 11 provides drugscomprising the agent for protecting and promoting liver function claimedin one of claims 8-10 as an active ingredient for protecting andpromoting liver function.

[0079] The drugs of the present invention comprise periandrins,Brazilian licorice extract, or one or more constituents contained in theextract. Thus, they have an effect of protecting and promoting liverfunction.

[0080] Also, the drugs of the present invention inhibit cytokineproduction by protecting and promoting liver function and ease symptomsof inflammation, allergy, atopy, and pollinosis, for example.

[0081] The drugs of the present invention are also advantageous in thatthey have no harmful side effect.

[0082] The drugs of the present invention, as well as the drugs claimedin claim 4, can be made into various forms and compound otheringredients.

[0083] (12) The invention set forth in claim 12 provides foodscomprising the agent for protecting and promoting liver function claimedin one of claims 8-10 as an active ingredient for protecting andpromoting liver function.

[0084] The foods of the present invention comprise periandrins,Brazilian licorice extract, or one or more constituents contained in theextract. Thus, they have effect of protecting and promoting liverfunction.

[0085] Also, the foods of the present invention inhibit cytokineproduction by protecting and promoting liver function and ease symptomsof inflammation, allergy, atopy, and pollinosis, for example.

[0086] Accordingly, an intake of the foods of the present invention canprotect and promote liver function and improve various inflammatorysymptoms by atopy, pollinosis, etc., for example.

[0087] The foods of the present invention are also advantageous in thatthey have no harmful side effect.

[0088] The foods of the present invention, as well as the foods claimedin claim 6, can be made into various forms and compound otheringredients.

[0089] (13) The invention set forth in claim 13 provides food materialscomprising the agent for protecting and promoting liver function claimedin one of claims 8-10 as an active ingredient for protecting andpromoting liver function.

[0090] The food materials of the present invention comprise periandrins,Brazilian licorice extract, or one or more constituents contained in theextract. Thus, they have an effect of protecting and promoting liverfunction.

[0091] Also, the food materials of the present invention inhibitcytokine production by protecting and promoting liver function and easesymptoms of inflammation, allergy, atopy, and pollinosis, for example.

[0092] Accordingly, an intake of foods compounding the food materials ofthe present invention can protect and promote liver function and improvevarious inflammatory symptoms by atopy, pollinosis, etc., for example.

[0093] The foods of the present invention are also advantageous in thatthey have no harmful side effect.

[0094] The food materials of the present invention can be a sweetener,for example.

[0095] The food materials of the present invention, as well as the foodmaterials claimed in claim 7, can compound various ingredients and canbe added to various foods.

[0096] (14) The invention set forth in claim 14 provides ananti-inflammatory agent comprising periandrins as an active ingredient.

[0097] The anti-inflammatory agent of the present invention compriseperiandrins, and thus, it shows an anti-inflammatory effect.

[0098] Particularly, the anti-inflammatory agent of the presentinvention can reduce inflammation due to autoimmune diseases (such asrheumatism (rheumatoid arthritis, for example), systemic lupuserythematosus, couagenosis, multiple sclerosis, and atopies) byinhibiting cytokine production.

[0099] The anti-inflammatory agent of the present invention is alsoadvantageous in that they have no harmful side effect. Therefore, it canbe administered over a long period in treatment of autoimmune diseasesand after organ transplant, for example, without developing any sideeffect unlike the conventional steroids.

[0100] The anti-inflammatory agent of the present invention can compoundperiandrins separated and purified from Brazilian licorice extract, forexample. In this case, since polysaccharide contained in Brazilianlicorice extract is not entered into the agent for protecting andpromoting liver function, it is difficult for the agent for protectingand promoting liver function to get moldy.

[0101] Examples of the aforementioned periandrins are periandrin I(PD-I), periandrins II (PD-II), periandrin III (PD-III), periandrin IV(PD-IV), etc.

[0102] (15) The invention set forth in claim 15 provides ananti-inflammatory agent comprising one of Brazilian licorice extract andone or more constituents contained in the extract as an activeingredient.

[0103] The anti-inflammatory agent of the present invention comprisesthe extract, or one or more constituents contained in the same, andthus, it shows an anti-inflammatory effect.

[0104] Particularly, the anti-inflammatory agent of the presentinvention can reduce inflammation due to autoimmune diseases (such asrheumatism (rheumatoid arthritis, for example), systemic lupuserythematosus, collagenosis, multiple sclerosis, and atopies) byinhibiting cytokine production.

[0105] The anti-inflammatory agent of the present invention is alsoadvantageous in that they have no harmful side effect. Therefore, it canbe administered over a long period in treatment of autoimmune diseasesand after organ transplant, for example, without developing any sideeffect unlike the conventional steroids.

[0106] Examples of the aforementioned one or more constituents aretriterpenoid glycosides and triterpenoid aglycones, such as periandrinsI, II, III, IV and V, periandradulcins A, B and C, and aglycones of theaforementioned. In short, they are triterpenoid glycosides and/ortriterpenoid aglycones contained in Brazilian licorice extract.

[0107] (16) The invention set forth in claim 16 provides theanti-inflammatory agent claimed in claim 15, one or more constituents ofthe aforementioned comprise periandrins.

[0108] The anti-inflammatory agent of the present invention comprisesperiandrins contained in Brazilian licorice extract, and thus, it showsan anti-inflammatory effect.

[0109] Also, the anti-inflammatory agent of the present invention, aswell as the anti-inflammatory agent claimed in claim 14, can reduceinflammation due to autoimmune diseases (such as rheumatism (rheumatoidarthritis, for example), systemic lupus erythematosus, collagenosis,multiple sclerosis, and atopies) by inhibiting cytokine production.

[0110] The anti-inflammatory agent of the present invention is alsoadvantageous in that they have no harmful side effect. Therefore, it canbe administered over a long period in treatment of autoimmune diseasesand after organ transplant, for example, without developing any sideeffect unlike the conventional steroids.

[0111] (17) The invention set forth in claim 17 provides animmunosuppressant comprising periandrins as an active ingredient.

[0112] Also, the immunosuppressant, comprising periandrins, of thepresent invention can ease symptoms of autoimmune diseases (caused byabnormality in human immune system mistaking cells or tissues of itselffor antigens due to genetic predisposition and low molecular extrinsicfactor called hapten (such as, rheumatism (rheumatoid arthritis, forexample), systemic lupus erythematosus, collagenosis, multiplesclerosis, and atopies).

[0113] Specifically, this autoimmune agent is unlikely to causeaggravation (i.e. rebound) of the symptoms after the administration isstopped.

[0114] Additionally, since the autoimmune agent of the presentinvention, as well as the anti-inflammatory agent claimed in claim 14,has no harmful side effect, it can be administered over a long period.

[0115] The autoimmune agent of the present invention can compoundperiandrins separated and purified from Brazilian licorice extract, forexample. In this case, since polysaccharide contained in Brazilianlicorice extract is not entered into the agent for protecting andpromoting liver function, it is difficult for the agent for protectingand promoting liver function to get moldy.

[0116] Examples of the aforementioned periandrins are periandrin I(PD-I), periandrins II (PD-II), periandrin III (PD-III), periandrin IV(PD-IV), etc.

[0117] (18) The invention set forth in claim 18 provides animmunosuppressant comprising one of Brazilian licorice extract and oneor more constituents contained in the extract as an active ingredient.

[0118] The immunosuppressant, comprising one of Brazilian licoriceextract and one or more constituents contained in the extract, of thepresent invention can ease symptoms of autoimmune diseases (caused byabnormality in human immune system mistaking cells or tissues of itselffor antigens due to genetic predisposition and low molecular extrinsicfactor called hapten (such as, rheumatism (rheumatoid arthritis, forexample), systemic lupus erythematosus, collagenosis, multiplesclerosis, and atopies).

[0119] Also, the autoimmune agent of the present invention has noharmful side effect, and thus it can be administered over a long period.

[0120] Examples of the aforementioned one or more constituents aretriterpenoid glycosides and triterpenoid aglycones, such as periandrinsI, II, III, IV and V, periandradulcins A, B and C, and aglycones of theaforementioned. In short, they are triterpenoid glycosides and/ortriterpenoid aglycones comprised in Brazilian licorice extract.

[0121] (19) The invention set forth in claim 19 provides the autoimmuneagent claimed in claim 18, one or more constituents of theaforementioned comprise periandrins.

[0122] The immunosuppressant, comprising periandrins as one or moreconstituents contained in Brazilian licorice extract, of the presentinvention can ease symptoms of autoimmune diseases (caused byabnormality in human immune system mistaking cells or tissues of itselffor antigens due to genetic predisposition and low molecular extrinsicfactor called hapten (for example, rheumatism (such as rheumatoidarthritis), systemic lupus erythematosus, collagenosis, multiplesclerosis, and atopies).

[0123] Also, the autoimmune agent of the present invention, as well asthe autoimmune agent claimed in claim 17, has no harmful side effect,and thus it can be administered over a long period.

[0124] (20) The invention set forth in claim 20 provides drugscomprising the anti-inflammatory agent claimed in one of claims 12-14 asan anti-inflammatory active ingredient.

[0125] The drugs of the present invention comprise periandrins,Brazilian licorice extract, or one or more constituents contained in theextract. Thus, they have an anti-inflammatory effect.

[0126] Particularly, the drugs of the present invention reduceinflammation due to autoimmune diseases (such as rheumatism (rheumatoidarthritis, for example), systemic lupus erythematosus, collagenosis,multiple sclerosis, and atopies).

[0127] The anti-inflammatory agent of the present invention is alsoadvantageous in that it has no harmful side effect.

[0128] The drugs of the present invention, as well as the drugs claimedin claim 4, can be made into various forms and compound otheringredients.

[0129] (21) The invention set forth in claim 21 provides cosmeticscontaining the anti-inflammatory agent claimed in one of claims 14-16,as an anti-inflammatory active ingredient.

[0130] The cosmetics of the present invention comprise periandrins,Brazilian licorice extract, or one or more constituents of the extract.Therefore, they have an anti-inflammatory effect (i.e. effect by whichinflammation due to autoimmune diseases such as rheumatism (rheumatoidarthritis, for example), systemic lupus erythematosus, collagenosis,multiple sclerosis, and atopies is reduced).

[0131] Accordingly, use of the cosmetics of the present invention canreduce inflammation caused by the aforementioned symptoms.

[0132] The cosmetics of the present invention are also advantageous inthat they have no side effect.

[0133] The cosmetics of the present invention, as well as the cosmeticsclaimed in claim 5, can be made into various forms and products.

[0134] (22) The invention set forth in claim 22 provides foodscomprising the anti-inflammatory agent claimed in one of claims 14-16 asan anti-inflammatory active ingredient.

[0135] The foods of the present invention comprise periandrins,Brazilian licorice extract, or one or more constituents contained in theextract. Therefore, they have an anti-inflammatory effect (i.e. effectby which inflammation due to autoimmune diseases such as rheumatism(rheumatoid arthritis, for example), systemic lupus erythematosus,collagenosis, multiple sclerosis, atopies, etc. is reduced).

[0136] Accordingly, an intake of the foods of the present invention, forexample, can reduce inflammation due to the aforementioned symptoms.

[0137] The foods of the present invention are also advantageous in thatthey have no harmful side effect.

[0138] The foods of the present invention, as well as the foods claimedin claim 6, can be made into various forms and compound otheringredients.

[0139] (23) The invention set forth in claim 23 provides food materialscomprising the anti-inflammatory agent claimed in one of claims 14-16 asan anti-inflammatory active ingredient.

[0140] The food materials of the present invention comprise periandrins,Brazilian licorice extract, or one or more constituents contained in theextract. Therefore, they have an anti-inflammatory effect (i.e. effectby which inflammation due to autoimmune diseases such as rheumatism(rheumatoid arthritis, for example), systemic lupus erythematosus,collagenosis, multiple sclerosis, atopies, etc. is reduced).

[0141] Accordingly, an intake of the food materials of the presentinvention, for example, can reduce inflammation due to theaforementioned symptoms.

[0142] The food materials of the present invention can be a sweetener,for example.

[0143] The food materials of the present invention, as well as the foodmaterials claimed in claim 7, can compound various ingredients and canbe added to various foods.

[0144] (24) The invention set forth in claim 24 provides drugscomprising the immunosuppressant claimed in one of claims 17-19 as animmunosuppressive active ingredient.

[0145] The drugs of the present invention comprise periandrins,Brazilian licorice extract, or one or more constituents contained in theextract. Thus, they have an immunosuppressive effect.

[0146] Accordingly, the drugs of the present invention reducedevelopment of autoimmune diseases (such as rheumatism (rheumatoidarthritis, for example), systemic lupus erythematosus, collagenosis,multiple sclerosis, and atopies) and also ease the symptoms thereof.

[0147] The immunosuppressant of the present invention is alsoadvantageous in that it has no harmful side effect.

[0148] The drugs of the present invention, as well as the drugs claimedin claim 4, can be made into various forms and compound otheringredients.

[0149] (25) The invention set forth in claim 25 provides cosmeticscomprising the immunosuppressant claimed in one of claims 15-17, as animmunosuppressive active ingredient.

[0150] The cosmetics of the present invention comprise periandrins,Brazilian licorice extract, or one or more constituents of the extract.Therefore, they have an immunosuppressive effect.

[0151] Accordingly, use of the cosmetics of the present invention canreduce development of autoimmune diseases such as rheumatism (rheumatoidarthritis, for example), systemic lupus erythematosus, collagenosis,multiple sclerosis, and atopies, and ease the symptoms thereof.

[0152] The cosmetics of the present invention are also advantageous inthat they have no side effect.

[0153] The cosmetics of the present invention, as well as the cosmeticsclaimed in claim 5, can be made into various forms and products.

[0154] (26) The invention set forth in claim 26 provides foodscomprising the immunosuppressant claimed in one of claims 15-17 as animmunosuppressive active ingredient.

[0155] The foods of the present invention comprise periandrins,Brazilian licorice extract, or one or more constituents contained in theextract. Therefore, they have an immunosuppressive effect.

[0156] Accordingly, an intake of the foods of the present invention, forexample, can reduce development of autoimmune diseases such asrheumatism (rheumatoid arthritis, for example), systemic lupuserythematosus, collagenosis, multiple sclerosis, atopies) and ease thesymptoms thereof.

[0157] The foods of the present invention are also advantageous in thatthey have no harmful side effect.

[0158] The foods of the present invention, as well as the foods claimedin claim 6, can be made into various forms and compound otheringredients.

[0159] (27) The invention set forth in claim 27 provides food materialscomprising the immunosuppressant claimed in one of claims 15-17 as animmunosuppressive active ingredient.

[0160] The food materials of the present invention comprise periandrins,Brazilian licorice extract, or one or more constituents contained in theextract. Therefore, they have an immunosuppressive effect.

[0161] Accordingly, an intake of the food materials of the presentinvention, for example, can reduce development of autoimmune diseasessuch as rheumatism (rheumatoid arthritis, for example), systemic lupuserythematosus, collagenosis, multiple sclerosis, atopies)

[0162] and ease the symptoms thereof.

[0163] The food materials of the present invention can be a sweetener,for example.

[0164] The food materials of the present invention, as well as the foodmaterials claimed in claim 7, can compound various ingredients and canbe added to various foods.

BRIEF DESCRIPTION OF THE DRAWINGS

[0165]FIG. 1 is an explanatory diagram of an experimentation methodaccording to an experiment 1;

[0166]FIG. 2 is an explanatory diagram showing weight changes of mice inan experiment 2;

[0167]FIG. 3 is an explanatory diagram showing changes in evaluationvalues of arthritis in the experiment 2;

[0168]FIG. 4 is an explanatory diagram showing changes in foot padswelling in the experiment 2;

[0169]FIG. 5 is an explanatory diagram showing changes in evaluationvalues of encephalomyelitis in an experiment 3;

[0170]FIG. 6 is an explanatory diagram showing weight changes of mice inan experiment 4;

[0171]FIG. 7 is an explanatory diagram showing changes in evaluationvalues of encephalomyelitis in the experiment 4; and

[0172]FIG. 8 is an explanatory diagram contrastively showing an effectof inhibiting hydroxysteroid dehydrogenase activation as a side effectin an experiment 5.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT

[0173] Cytokine production inhibitors, agents for protecting andpromoting liver function, anti-inflammatory agents, immunosuppressants,drugs, cosmetics, foods, and food materials according to the presentinvention will now be explained below.

[0174] a) Firstly, extract, etc. obtained in examples and comparativeexamples are described.

EXAMPLE 1

[0175] Brazilian licorice extract was obtained using the following stepsI-V. These steps were repeated three times.

[0176] I. 300 g of dried rootstalk of Brazilian licorice was ground, andthen, extraction was performed for an hour with 900 g of 30 wt % ethanolat the circumfluent temperature.

[0177] II. The liquid extract obtained in step I was suctioned andfiltered, and then, solid-liquid separation was performed.

[0178] III. The solid material obtained in step II was again ground andextraction was performed as in step I, and then the obtained liquidextract was separated as in step II.

[0179] IV. The solid material obtained in step III was again ground andextraction was performed as in step I, and then the obtained liquidextract was separated as in step II.

[0180] V. All the liquid extracts obtained in steps I, III and IV werecollected together, vacuumed and concentrated, and then dried byspray-drying to obtain solid extract.

[0181] Average yield and standard deviation of the last extract obtainedin the above manner were, respectively, 1.03 g and 0.49 g.

[0182] This Brazilian licorice extract, as can be understood from thelater described experiments, has effects of inhibiting cytokineproduction and protecting and promoting liver function. Accordingly,this Brazilian licorice extract can be used as cytokine productioninhibitors, agents for protecting and promoting liver function, drugs,cosmetics, foods, and food materials.

EXAMPLE 2

[0183] The extraction method of Example 1 was exercised in basically thesame manner to obtain Brazilian licorice extract. However, in this case,900 g of 20 wt % ethanol was used in place of 900 g of 30 wt % ethanolfor extraction.

[0184] Average yield and standard deviation of the extract obtained inthe above manner were, respectively, 1.53 g and 2.11 g.

[0185] This Brazilian licorice extract, as can be understood from thelater described experiments, has an anti-inflammatory effect and animmunosuppressive effect. Accordingly, this Brazilian licorice extractcan be used as anti-inflammatory agents, immunosuppressants, drugs,cosmetics, foods, and food materials.

EXAMPLE 3

[0186] Brazilian licorice extract obtained in Example 2 was fractionatedusing the following steps I-IV.

[0187] I. 575 g of Brazilian licorice extract was dissolved in a 20 v/v% methanol solution, and introduced into a column (300×400 mm) which wasfilled with ion-exchange resin (DIAION HP-20, produced by MitsubishiChemical Corporation), to be suctioned and cleaned with a 20 v/v %methanol solution.

[0188] II. The cleaning component obtained in step I was dried underreduced pressure, and 106 g of a solid content (BL. 1) was thusobtained.

[0189] III. The column is cleaned with a 50 v/v % methanol solution, andby drying under reduced pressure the resulted cleaning solution, 63.4 gof a solid content (BL. 2) was obtained.

[0190] IV. The column is cleaned with 80 v/v % methanol solution, and bydrying under reduced pressure the resulted cleaning solution, 296 g of asolid content (BL. 3) was obtained.

[0191] In order to prove that periandrins are contained in the BL. 3,the following method was used.

[0192] Particularly, purified periandrins obtained in later describedExperiment 4 were made a preparation. Periandrins were identifiedthrough silica gel thin-layer chromatography using two types ofdeveloping solvents (one of which contains chloroform, methanol, waterand acetic acid in the proportions of 70:35:5:5; and the other of whichcontains ethyl acetate, isopropyl alcohol, water, ethanol anddiethylamine in the proportions of 20:10:8:1:0.3).

[0193] The BL. 3 (which is a constituent contained in Brazilian licoriceextract and which contains periandrins) has an excellentimmunosuppressive effect as seen in the later explained Experiment 4.

EXAMPLE 4

[0194] From the extract obtained in Example 1 or 2, purified periandrins(PD-I, PD-II, PD-III and PD-IV) were obtained by way of the usualseparation purification method using column chromatography.

[0195] Nuclear magnetic resonance spectrum and mass spectrum of theobtained PD-I, PD-II, PD-III and PD-IV were measured respectively, andit was found that each of them is not a mixture.

[0196] The periandrins (PD-I, PD-II, PD-III and PD-IV) obtained inExample 4 are, as shown in later explained Experiment 5, hardly inhibitsactivity of 11 β-hydroxysteroid dehydrogenase.

[0197] Therefore, these periandrins do not develop pseudoaldosteronism,which is considered to be caused by inhibition of 11 β-hydroxysteroiddehydrogenase activity.

[0198] These penandrins have effects of inhibiting cytokine productionand protecting and promoting liver function, an anti-inflammatoryeffect, and an immunosuppressive effect.

EXAMPLE 5

[0199] From Brazilian licorice extract obtained in Example 2, purifiedperiandrins (PD-I, PD-II, PD-III and PD-IV) were obtained using thefollowing steps I-IV.

[0200] I. 4-nitrobenzylbromide (produced by Tokyo Kasei Kogyo Co.,Ltd.), dimethylformaldehyde as a solvent, and triethylamine as ahydrobromic scavenger were added to Brazilian licorice extract.According to the usual manner, 4-nitrobenzylization was performed.

[0201] As a result, 4-nitrobenzylized PD-I, PD-II, PD-III and PD-IV wereobtained.

[0202] II. After the 4-nitrobenzylization in step I,dimethylformaldehyde was vacuumed and distilled, to extract a reactionproduct containing 4-nitrobenzylized PD-I, PD-II, PD-III and PD-IV withethyl acetate.

[0203] III. From the ethyl acetate extract obtained in step II, the4-nitrobenzylized PD-I, PD-II, PD-III and PD-IV were separated andpurified through chromatography with silica gel (WAKOGEL C-200, producedby Wako Pure Chemical Industries,Ltd.) and octadecylsilane (produced byChromatorex, Fuji Silysia Chemical Ltd.).

[0204] IV. The 4-nitrobenzylized PD-I, PD-II, PD-III and PD-IV obtainedin step II were dissolved in an acetic solvent, and then, by adding zincpowder, denitrobenzilized reaction was performed to obtain purifiedperiandrins (PD-I, PD-II, PD-III and PD-IV).

[0205] The periandrins obtained in Example 5 are, like the periandrinsobtained in Example 4, hardly inhibits activity of 11 β-hydroxysteroiddehydrogenase and thus do not develop pseudoaldosteronism.

[0206] Furthermore, the periandrins obtained in Example 4 or 5 have, ascan be clearly seen from later explained Experiment 6, effects ofinhibiting cytokine production and protecting and promoting liverfunction, an anti-inflammatory effect and an immunosuppressive effect.

COMPARATIVE EXAMPLE 1

[0207] Chinese licorice extract was obtained in the same manner as inthe aforementioned Example 1.

[0208] Chinese licorice is a typical species of licorice, which has beenused as an agent for protecting liver function.

[0209] Average yield and standard deviation of the extract obtained inthe above manner were, respectively, 27.0 g and 1.03 g.

COMPARATIVE EXAMPLE 2

[0210] Chinese licorice extract was obtained in the same manner as theaforementioned Example 2.

[0211] Average yield and standard deviation of the extract obtained inthe above manner were, respectively, 28.5 g and 2.18 g.

[0212] b) Secondly, experiments conducted to confirm the effects of theextract obtained in the aforementioned Examples are explained.

Experiment 1

[0213] The following experiment was performed to examine the effects ofinhibiting cytokine production and lowering GOT activity of the extractsobtained in Example 1 and Comparative Example 1. The experimentprocedures are shown in FIG. 1.

[0214] I. Preparation of mice to be used in the experiment

[0215] Male mice of ddy system (Japan SLC, Inc.), each of which weighs18-20 g, are prepared for the experiment. The mice are divided into sixgroups, namely, A, B, C, D, E and F, each consisting of eight mice.

[0216] II. Injection of Propionivacterium acnes suspension

[0217] 5 mg of Propionivacterium acnes suspension was injected into thevein of the aforementioned mice respectively, to cause hepatopathy.

[0218] III. Injection of Brazilian licorice extract and comparativesample

[0219] A solvent without drugs was injected into the mice in group A.

[0220] Brazilian licorice extract obtained in Example 1 was injectedinto the abdominal cavity of the mice in groups B and C four times intotal, that is, one day, three days, five days and seven days after theinjection of the propionivacterium acnes suspension in step II. Thedosage per one injection was 25 mg, respectively.

[0221] Chinese licorice extract obtained in Comparative Example 1 wasinjected into the abdominal cavity of the mice in groups D and E fourtimes in total, that is, one day, three days, five days and seven daysafter the injection of the propionivacterium acnes suspension in stepII. The dosage per one injection was 25 mg, respectively.

[0222] 5 mg of prednisolone acetate was injected into the abdominalcavity of the respective mice in group F four times in total, that is,one day, three days, five days and seven days after the injection of thepropionivacterium acnes suspension in step II.

[0223] Prednisolone acetate is a typical type of prednisolone, which hasbeen used as a cytokine production inhibitor.

[0224] IV. Sampling of blood

[0225] 10 μg of lipopolysaccharide was injected into the vein of themice in all groups respectively, thirty minutes after the fourthinjection in step II (as to the mice in group A, the injection wasperformed at the same timing as the mice in the other groups). Then, twohours later, blood was taken from all the mice to obtain serum. Therewas no visible change in weight and food and water consumption of themice.

[0226] V. Measurement of cytokine in blood

[0227] From the serum obtained in step III, measurement of interleukin-6(IL-6), which is a kind of cytokine, was performed.

[0228] For the measurement, a commercially available measurement kitwhich adopts Fluorescence Linked immuno-Sorbent Assay using fluorescencelabeling antibody was used.

[0229] VI. Measurement of GOT in blood

[0230] From the serum obtained in step III, glutamic-oxaloacetictransaminase activity (GOT) was measured.

[0231] For the measurement, a commercially available measurement kitwhich adopts Enzyme Linked immuno-Sorbent Assay using enzyme labelingantibody was used.

[0232] Blood cytokine (IL-6) levels of the respective eight mice weremeasured per each of the groups A-F. Table 1 shows the average yield andstandard deviation.

[0233] The measured values in Table 1 show IL-6 levels (unit: picogram)per 1 ml serum. TABLE 1 Average of Standard Group eight mice deviation A360349 93784 B 40845 11348 C 76772 58125 D 280986 97265 E 191646 91081 F26473 15300

[0234] As shown in Table 1, the blood IL-6 levels of the mice in groupsB and C to which Brazilian licorice extract was injected were remarkablyreduced compared to the blood IL-6 levels of the mice in group A withoutdrug injection and of the mice in groups D and E to which Chineselicorice extract was injected. The levels were rather close to the bloodIL-6 levels of group F to which prednisolone acetate was injected.

[0235] In view of the above, it is clear that Brazilian licorice extracthas an effect of inhibiting cytolkine production which equals to theeffect by prednisolone acetate.

[0236] Table 2 shows average yield and standard deviation of blood GOTactivity levels of the respective eight mice measured per each of thegroups A-F.

[0237] The measured values in Table 2 are shown in Kermen Units. TABLE 2Average of Standard Group eight mice deviation A 918.160 256.863 B244.359 31.405 C 201.840 18.893 D 462.386 118.867 E 412.544 159.981 F350.705 68.166

[0238] As can be seen in Table 2, the blood GOT activity levels of themice in groups B and C to which Brazilian licorice extract was injectedwere lower than the blood GOT activity levels of any other groups.

[0239] In view of the above, Brazilian licorice extract has a superioreffect of protecting and promoting liver function to Chinese licoriceextract or prednisolone acetate.

Experiment 2

[0240] Anti-inflammatory effect and immunosuppressive effect of theextract obtained in Example 2 and Comparative Example 2 were examined bymeans of collagen induction arthritis model experiment.

[0241] I. Preparation of mice to be used in experiment

[0242] DBJ/1J mice were used in the experiment. The mice are dividedinto six groups, namely, G, H, I, J, K and L, each consisting of sixmice.

[0243] II. Induction of arthritis by injection of emulsion to mice(immunity)

[0244] An equal amount of 8 mg/ml concentration of a collagen solution(the solvent is phosphate buffered saline containing 0.01 mol/L aceticacid) and fetal bovine serum containing 4 mg/ml concentration ofsupersonically treated Microbacterium tuberculosis H37Ra were mixed toprepare emulsion.

[0245] 50 μl (200 μg/head) of the emulsion was then injected to the backneck skin of each of the mice under ether (first immunity).

[0246] Three weeks after the first immunity, 50 μl (200 μg/head) of theemulsion was injected to the tail head skin of each of the mice (secondimmunity).

[0247] Arthritis was induced in the respective mice due to the first andsecond immunities.

[0248] III. Injection of Brazilian licorice extract and comparativesample to mice

[0249] A solvent without drugs was injected into the mice in group G.

[0250] Brazilian licorice extract obtained in Example 2 was diluted byphysiological saline so that the dosage is 6 mg/head, and the obtaineddiluted solution was injected into the abdominal cavity of the mice ingroup H by 1 ml every other day after the first immunity.

[0251] Brazilian licorice extract obtained in Example 2 was diluted byphysiological saline so that the dosage is 12.5 mg/head, and theobtained diluted solution was injected into the abdominal cavity of themice in group I by 1 ml every other day after the first immunity.

[0252] Chinese licorice extract obtained in Comparative Example 2 wasdiluted by physiological saline so that the dosage is 6 mg/head, and theobtained diluted solution was injected into the abdominal cavity of themice in group J by 1 mg every other day after the first immunity.

[0253] Chinese licorice extract obtained in Comparative Example 2 wasdiluted by physiological saline so that the dosage is 12.5 mg/head, andthe obtained diluted solution was injected into the abdominal cavity ofthe mice in group K by 1 mg every other day after the first immunity.

[0254] Prednisolone acetate (produced by Shionogi & Co., Ltd.) which isa control drug was suspended by physiological saline so that the dosageis 5 mg/Kg of body weight, and the obtained suspended solution wasinjected into the abdominal cavity of each mouse in group L by 1 mlevery other day after the first immunity.

[0255] The above injections were performed all through the test period(for eight weeks after the first immunity).

[0256] IV. Weight measurement of mice

[0257] The weight of the mice in the respective groups were measuredevery day after the first immunity and the average weight per each ofthe groups was calculated. The results are shown in FIG. 2.

[0258] As can be seen from FIG. 2, there was no visible reduction ofaverage weight in the mice of groups H and I to which Brazilian licoriceextract was injected. In addition, there was no reduction in averageweight in the mice of groups H and I after the second immunity, either.

[0259] On the other hand, the average weights of the mice in group Gwithout drug injection, groups J and K to which Chinese licorice extractwas injected, and group K to which prednisolone acetate was injectedwere reduced significantly compared to the average weights of mice ingroups H and I.

[0260] V. Accordingly, Brazilian licorice extract hardly causes sideeffects and is highly secure compared to Chinese licorice extract andprednisolone acetate.

[0261] Evaluation of Arthritis Condition (Arthritis Index)

[0262] Clinical symptoms of arthritis of mice in the respective groupswere evaluated by one and the same observer using 6 rankings from zeroto five (the larger the value is, the worse the symptom is), and theaverages per the respective groups were calculated. These evaluationswere performed in 0, 3^(rd), 4^(th), 5^(th), 6^(th), 7^(th) and 8^(th)week after the first immunity The results were shown in FIG. 3 and Table3. In Table 3, values in the upper section show the average in therespective groups, and the values in the lower section show the standarddeviation. TABLE 3 Primary Drug type Gr. Dosage immunity 3rd week 4thweek 5th week 6th week 7th week 8th week None G — 0 0.688 1.313 3.1252.688 2.688 2.688 0 0.377 0.4 0.611 0.582 0.49 0.462 B.L. H   6 mg/ 0 00.25 1.625 1.625 0.938 0.929 head 0 0 0.189 0.653 0.673 0.29 0.254 I12.5 mg/ 0 0 0 0.9 1.2 0.8 0.6 head 0 0 0 0.9 0.846 0.339 0.367 C.L. J  6 mg/ 0 0 0.5 2.833 2.75 2.583 2.167 head 0 0 0.5 0.843 0.588 0.3960.307 K 12.5 mg/ 0 0 0.571 3 3.071 3.071 2.357 head 0 0 0.277 0.5560.539 0.297 0.404 Pred. L   5 mg/ 0 0 0 0 0 0.125 0.125 head 0 0 0 0 00.125 0.125

[0263] As can be seen from FIG. 3 and Table 3, the arthritis symptoms ofmice in groups to which Brazilian licorice extract was injected (groupsH and I) were improved compared to the mice in group G without druginjection and groups J and K to which Chinese licorice extract wasinjected.

[0264] In view of the above, it is clear that Brazilian licorice extracthas superior anti-inflammatory and immunosuppressive effects to Chineselicorice extract.

[0265] As to the mice in groups H and I to which Brazilian licoriceextract was injected, the symptoms of the mice in group I to which largequantity (12.5 mg/head) of the extract was injected per one time werebetter improved than the mice in group H to which small quantity (6mg/head) of the extract was injected per one time. In short, the effectof improving arthritis symptoms by Brazilian licorice extract showeddose dependency.

[0266] VI. Measurement of foot pad swelling (foot pad volume)

[0267] The left and right hind foot pad volumes of the mice in therespective groups were measured using a plethysmometer (TK-101 UNICOM),and the average per each of the groups was calculated. These evaluationswere performed in 0, 3^(rd), 4^(th), 5^(th), 6^(th), 7^(th) and 8^(th)weeks after the first immunity. The results were shown in FIG. 4 andTable 4. The values in FIG. 4 are relative values based on the initialhind foot pad volumes. In Table 4, the values in the upper section showthe averages of the respective groups and the values in the lowersection show the standard deviation. TABLE 4 Primary Drug type Gr.Dosage immunity 3rd week 4th week 5th week 6th week 7th week 8th weekNone M — 100 112.9 111 143.3 141.2 124.7 122.7 0 4.517 6.084 6.821 5.5343.109 2.241 B.L. N   6 mg/ 100 107.9 104 123.1 126.4 114 114.7 head 01.34 1.871 6.793 5.543 3.117 2.509 O 12.5 mg/ 100 107.3 101 117.8 121.7116 112.4 head 0 3.588 2.527 6.113 6.04 3.417 2.478 C.L. P   6 mg/ 100100.9 100.7 130 148.6 124.6 123.4 head 0 3.644 4.638 12.752 8.55 4.3384.846 Q 12.5 mg/ 100 105.4 101.5 150.8 130.7 122.2 124.4 head 0 1.2873.145 7.663 6.619 4.528 4.313 Pred. R   5 mg/ 100 99.2 95.2 101.1 102.495.5 97.9 head 0 2.328 2.037 2.347 1.866 1.615 1.692

[0268] It is clear from FIG. 4 and Table 4 that there was a superioreffect of inhibiting foot pad swelling in the mice in groups H and I towhich Brazilian licorice extract was injected compared to the mice ingroup G without drug injection and groups J and K to which Chineselicorice extract was injected.

[0269] It should be noted that a superior effect of inhibiting foot padswelling was seen even in the mice in group I to which less Brazilianlicorice extract (6 mg/head) was injected compared to the mice in groupsG, J and K.

[0270] In view of the above, Brazilian licorice extract has superioranti-inflammatory and immunosuppressive effects to Chinese licoriceextract.

Experiment 3

[0271] Anti-inflammatory effect and immunosuppressive effect of theextract obtained in Example 2 and Comparative Example 2 were examined byautoimmune encephalomyelitis model experiment using rats.

[0272] I. Preparation of rats to be used in experiment

[0273] Eight weeks old male rats of DA system were used in theexperiment. The rats were divided into 6 groups, namely, M, N, O, P, Qand R, each consisting of six rats.

[0274] II. Induction of autoimmune encephalomyelitis by injection ofemulsion to rats

[0275] An equal amount of 2 mg/ml concentration of a bovine MyelineBasic Protein (MBP) solution (the solvent is phosphate buffered saline)and fetal bovine serum containing supersonically treated 4 mg/mlMicrobacterium tuberculosis H37Ra were mixed to prepare emulsion.

[0276] 50 μl of the emulsion was then injected hypodermically to hindleg foot pads of the respective rats under ether (in other words, 100 μl(100 μgMBP/head) was injected per rat), to induce autoimmuneencephalomyelitis.

[0277] III. Injection of Brazilian licorice extract and comparativesample

[0278] A solvent without drugs was injected into the rats in group M.

[0279] Brazilian licorice extract obtained in Example 2 was diluted byphysiological saline so that the dosage is 25 mg/head, and the obtaineddiluted solution was injected into the abdominal cavity of the rats ingroup N by 1 ml once a day for 12 consecutive days after the injectionof emulsion.

[0280] Brazilian licorice extract obtained in Example 2 was diluted byphysiological saline so that the dosage is 50 mg/head, and the obtaineddiluted solution was injected into the abdominal cavity of the rats ingroup O by 1 ml once a day for 12 consecutive days after the injectionof emulsion.

[0281] Chinese licorice extract obtained in Comparative Example 2 wasdiluted by physiological saline so that the dosage is 25 mg/head, andthe obtained diluted solution was injected into the abdominal cavity ofthe rats in, group P by 1 ml once a day for 12 consecutive days afterthe injection of emulsion.

[0282] Chinese licorice extract obtained in Comparative Example 2 wasdiluted by physiological saline so that the dosage is 50 mg/head, andthe obtained diluted solution was injected into the abdominal cavity ofthe rats in group Q by 1 ml once a day for 12 consecutive days after theinjection of emulsion.

[0283] Prednisolone acetate (produced by Shionogi & Co., Ltd.) which isa control drug was suspended by physiological saline so that the dosageis 5 mg/Kg of body weight, and the obtained suspended solution wasinjected into the abdominal cavity of each rat in group R by 1 ml once aday for 12 consecutive days after the injection of emulsion.

[0284] IV. Evaluation of encephalomyelitis (Clinical score)

[0285] After the injection of emulsion, the rats in the respectivegroups were observed every day to evaluate symptoms of encephalomyelitis(Clinical sore). The symptoms were evaluated by six rankings from zeroto five (the larger the value, the worse the symptom is) according tothe criteria shown in Table 5, and the averages per the respectivegroups were calculated. The above observations were conducted for 25consecutive days after the injection of emulsion. TABLE 5 Value Criteriaof clinical score of encephalomyelitis 0 No change 1 Paralysis of tail 2Incomplete paralysis of hind legs 3 Complete paralysis of hind legs andincomplete paralysis of front legs 4 Paralysis of limbs, incontinence 5Dead

[0286] The results of the evaluation are shown in FIG. 5. As can beclearly seen from FIG. 5, a few rats in group N to which 25 mg/head ofBrazilian licorice extract was injected developed encephalomyelitisduring and after the injections. However, most of the rats did notdevelop encephalomyelitis.

[0287] The mice in group O to which 50 mg/head of Brazilian licoriceextract was injected did not develop encephalomyelitis during theinjections at all. However, they developed encephalomyelitis after theinjections were completed.

[0288] On the other hand, the rats in group M without drug injection wasand groups P and Q to which Chinese licorice extract was injecteddeveloped encephalomyelitis 7-8 days after the injection of emulsion.The symptoms reached its peak on 11^(th)-12^(th) days and weredisappeared on 14^(th) day. The symptoms of the rats in group Q to whichmore Chinese licorice extract was injected (50 mg/head) were slightlybetter than the rats in groups M and P.

[0289] A few rats in group R to which prednisolone acetate was injecteddeveloped encephalomyelitis 8-11 days after the injection of emulsion.

[0290] In view of the above, Brazilian licorice extract has superioranti-inflammatory and immunosuppressive effects to Chinese licoriceextract.

Experiment 4

[0291] The immunosuppressive effect of the BL. 1, BL. 2 and BL. 3, whichare fractionated components of Brazilian licorice extract obtained inExample 3, was examined by autoimmune encephalomyelitis model experimentusing rats.

[0292] The manner of experiment was basically the same as theaforementioned Experiment 3.

[0293] However, the number of groups of rats were reduced to five, oneof which included the rats to which the BL. 1 was injected, another ofwhich included the rats to which the BL. 2 was injected, another ofwhich included the rats to which the BL. 3 was injected, another ofwhich included the rats to which prednisolone acetate was injected, andanother of which included the rats without injection.

[0294] The dose per one injection of BL. 1, BL. 2 and BL. 3 was 6mg/head and in the form of physiological saline.

[0295] The dose per one injection of prednisolone acetate was 5 mg/Kg ofbody weight and in the form of physiological saline.

[0296]FIG. 6 shows weight changes of the rats in the respective groupsduring the experiment period.

[0297] The rats in the groups to which the BL. 1, BL. 2 and BL. 3 wereinjected gained about additional 5-10 g compared to the rats in thecontrol group on and after 8^(th) day after the immunity.

[0298] The rats in the group to which prednisolone acetate was injectedhad the least increase among the rats in the other groups.

[0299]FIG. 7 shows the evaluation results on encephalomyelitis. Theevaluation criteria are the same as the above Table 5.

[0300] Development of encephalomyelitis was strongly inhibited in therats to which the BL. 3 was injected compared to any of the rats in thegroups to which the BL. 1 and BL. 2 were injected, and the rats in thecontrol group. Development of encephalomyelitis was slightly inhibitedin the rats to which the BL. 2 was injected compared to the rats in thecontrol group.

[0301] In addition, there was no temporary development (for example,development caused on 15^(th)-16^(th) days in the rats of group O inExperiment 3) after completion of the injections in the rats in thegroup to which the BL. 3 was injected.

[0302] The results of Experiment 4 indicate that not the componentcontained in the BL. 1 (polar component such as polysaccharide which isnot held by suction chromatography carrier such as HP-20), but the polarcomponent such as triterpenoid glycoside (for example, periandrins)contained in the BL. 3 has an immunosuppressive effect.

Experiment 5

[0303] In order to evaluate the effect of inhibiting 11 βhydroxysteroiddehydrogenase activity by the purified periandrins (PD-I, PD-II, PD-III,PD-IV) obtained in Example 4 or 5, IC 50 values (concentration in whichthe 50 percent of activity of the above enzyme is inhibited) of thepurified periandrins were measured by the following steps I-VI.

[0304] Note that inhibition of 11 β-hydroxysteroid dehydrogenaseactivity is considered to be a cause of pseudoaldosteronism.

[0305] I. An enzymatic solution containing rat liver microsome as apreparation of 11 β-hydroxysteroid dehydrogenase was prepared as below.

[0306] Fresh rat liver was ice-cooled and homogenized in 0.25M sucrosesolution. Then, at the temperature of 4° C., it was centrifuged for 20minutes at 10,000×g.

[0307] After separating cell organelles of the nucleus, mitochondria andlysosome as deposits, the supernatant layer was further centrifuged at105,000×g for 60 minutes at the temperature of 4° C. to obtain amicrosome fraction as a precipitate.

[0308] The obtained microsome fraction was suspended by 0.25M sucrosesolution to produce an enzymatic solution, and the obtained solution wasstored at the temperature of −20 degrees till the time of measurement ofenzyme activity.

[0309] II. Measurement of enzyme activity

[0310] 1 mM of NADP (product of Oriental Yeast Co., Ltd,) as a coenzymeand an amount of PD-I having a predetermined concentration (ranging from2 to 1480 μM) as an inhibitor were added to 0.8 ml of Tris-HCI buffersolution having the concentration of 100 mM (PH8.0).

[0311] 0.2 ml of the enzymatic solution prepared in step I was added tothe obtained solution, and after three minutes of preincubation at 37°C., 0.02 ml of 5 mM hydrocortisone solution which was dissolved inmethanol was added to start a reaction between 11 β-hydroxysteroiddehydrogenase and PD-I. The reaction was continued for 30 minutes at thetemperature of 30° C., and then it was stopped by adding 0.5 ml of ethylacetate.

[0312] This solution was centrifuged and an ethyl acetate layer wasseparated into an Eppendorf tube. In the mean time, cortisone andsubstrate hydrocortisone generated by the reaction between 11β-hydroxysteroid dehydrogenase and substrate hydrocortisone aretransferred to the ethyl acetate layer.

[0313] Furthermore, the water phase after the centrifuge was againcentrifuged in the same manner as above by adding 0.5 ml of ethylacetate to separate an ethyl acetate layer. This process was repeatedtwo more times.

[0314] The ethyl acetate layers obtained by three time extraction wereput together into an Eppendorf tube and dried using a centrifugalevaporator.

[0315] III. Determination of generated cortisone

[0316] To prepare a measurement sample, 0.2 ml of methanol was addedinto the Eppendorf tube in which the ethyl acetate layers were dried instep II, and then the tube was left for a night so that cortisone wascompletely dissolved. Cortisone was determined using HPLC.

[0317] Particularly, quantity of cortisone contained in the sample wascalculated from a ratio of a peak area corresponding to cortisone whenthe measurement sample was introduced to HPLC to a peak area when 5 μlof standard solution (0.1 mg/ml of cortisone-methanol solution) wasintroduced.

[0318] The measurement conditions of HPLC were as follows. Stationaryphase: NovaPak cartridge C18, 3.9 × 150 mm (product of Japan Waters)Moving phase: 45% methanol solution containing 0.1% trifluoroacetic acid(isocratic) Flow rate: 0.5 ml/min Detection: UV 245 nm Injection amount:5 μl

[0319] IV. Relative activity (ratio of activity remained after reactionwith an inhibitor to the initial activity) of 11 β-hydroxysteroiddehydrogenase was calculated from the quantity of cortisone inaccordance with the following equation.

Relative activity=((C _(I) −C _(BL))/(C _(T) −C _(BL)))×100(%)

[0320] where

[0321] C_(T): cortisone quantity generated when no inhibitor is added;

[0322] C_(I): cortisone quantity generated when an inhibitor exists; and

[0323] C_(BL): cortisone quantity when enzyme is not added.

[0324] V. Measurement of relative activity when PD-II, PD-III, PD-IV,and glycirrhizin were added as an inhibitor instead of PD-I wasperformed in the same manner as in steps I-IV.

[0325] The concentration of the inhibitor PD-I, PD-II, PD-III or PD-IVin the measurement of relative activity is a predetermined value rangingfrom 2 to 1480 μM. The concentration of glycirrhizin is a predeterminedvalue ranging from 0.2 to 445 μM.

[0326] The system to which no inhibitor was added was considered as acontrol, and the system to which no substrate was added is considered asa blank.

[0327] VI. As shown in FIG. 6, relation between the concentrations ofthe respective inhibitors and the relative activity was plotted, and theinhibitor concentration of which relative activity is equal to 50% wasmade IC50 of the inhibitor.

[0328] As to the measurement results, on one hand, the IC50 value ofglycirrhizin was 26.2 μM, and on the other hand, the IC50 values ofPD-I, PD-II, PD-III and PD-IV were respectively 214 μM, 150 μM, 581 μM,240 μM, which were high in one order.

[0329] In other words, inhibitory activity of PD-I, PD-II, PD-III andPD-IV was about one fifth to one twentieth of glycirrhizin.

[0330] Accordingly, since the purified periandrins hardly inhibit 11β-hydroxysteroid dehydrogenase activity, pseudoaldosteronism, which isconsidered to be caused by inhibition of 11 β-hydroxysteroiddehydrogenase activity, is not developed by using the productscomprising the purified periandrins (cytokine production inhibitors,agents for protecting and promoting liver function, anti-inflammatoryagents, immunosuppressants, drugs, cosmetics, foods and food materials).

Experiment 6

[0331] The effects of inhibiting cytokine production and of protectingand promoting liver function with respect to PD-I, PD-II, PD-III andPD-IV obtained in Example 4 or 5 were examined in the same manner as inExperiment 1.

[0332] In this case, however, the number of groups of rats were madesix, one of which includes the rats to which PD-I was injected, anotherof which includes the rats to which PD-II was injected, another of whichincludes the rats to which PD-III was injected, another of whichincludes the rats to which PD-IV was injected, another of which includesthe rats to which prednisolone acetate was injected, and another ofwhich includes the rats to which a solvent without any drugs wasinjected.

[0333] The dose per one injection of PD-I, PD-II, PD-III and PD-IV was0.1-3 mg/Kg of body weight.

[0334] The dose per one injection of prednisolone acetate was 5 mg/Kg ofbody weight.

[0335] All of PD-I, PD-II, PD-III and PD-IV show the effects ofinhibiting cytokine production and protecting and promoting liverfunction. It was found that these components are at least one part ofthe active ingredient of Brazilian licorice extract.

[0336] Furthermore, the collagen induction arthritis model experimentand autoimmune encephalomyelitis model experiment shown in Experiments 2and 3 were applied using PD-I, PD-II, PD-III and PD-IV obtained inExperiment 4. As a result, these components eased the symptoms ofarthritis and encephalomyelitis like Brazilian licorice extract did.

[0337] From the above, the purified periandrins (PD-I, PD-II, PD-III andPD-IV) have anti-inflammatory effect and immunosuppressive effect.

[0338] The present invention should not be limited to the describedembodiments, and other modifications and variations might be possiblewithout departing from the scope of the invention.

[0339] For example, in the process of obtaining Brazilian licoriceextract, the solvent is not limited to ethanol, but other solvents (e.g.water, alcohols such as methanol, isopropanol, isobutanol, hexanol,methyl amyl alcohol, 2-ethyl butanol, n-octyl alcohol, etc., polyhydricalcohols and the derivatives such as glycerin, ethylene glycol, ethyleneglycol monomethyl ether, ethylene glycol monoethyl ether, propyleneglycol, propylene glycol monomethyl ether, propylene glycol monoethylether, triethylene glycol, 1,3-butylene glycol, hexylene glycol, etc.,ketones such as acetone, methyl ethyl ketone, methyl isobutyl ketone,etc., esters such as ethyl acetate, isopropyl acetate, etc., ethers suchas ethylether, isopropylether, n-bytylether, etc., aliphatichydrocarbons such as petroleum ether, n-hexane, n-pentane, n-butane,n-octane, cyclohexane, etc., and non polar solvents such astetrachloromethane, chloroform, trichloroethylene, benzol, toluene,etc.) can be used.

INDUSTRIAL AVAILABILITY

[0340] By adopting Brazilian licorice extract or periandrins as acytokine production inhibitor, it is possible to inhibit inflammation ofvarious diseases such as rheumatoid arthritis. Brazilian licoriceextract and periandrins can be used as an agent for protecting andpromoting liver function, an anti-inflammatory agent, and animmunosuppressant. Furthermore, foods, cosmetics, sweeteners and foodmaterials comprising Brazilian licorice extract also have the sameeffect as above.

What is claimed is:
 1. A cytokine production inhibitor comprisingperiandrins as an active ingredient.
 2. A cytokine production inhibitorcomprising one of Brazilian licorice extract and one or moreconstituents contained in the extract as an active ingredient
 3. Thecytokine production inhibitor as set forth in claim 2 wherein said oneor more constituents contain periandrins.
 4. Drugs comprising saidcytokine production inhibitor as set forth in one of claims 1-3 as ananti-inflammatory, anti-allergy, and anti-atopic active ingredient. 5.Cosmetics comprising said cytokine production inhibitor as set forth inone of claims 1-3 as an anti-inflammatory, anti-allergy, and anti-atopicactive ingredient.
 6. Foods comprising said cytokine productioninhibitor as set forth in one of claims 1-3 as an anti-inflammatory,anti-allergy, and anti-atopic active ingredient.
 7. Food materialscomprising said cytokine production inhibitor as set forth in one ofclaims 1-3 as an anti-inflammatory, anti-allergy, and anti-atopic activeingredient.
 8. An agent for protecting and promoting liver functioncomprising periandrins as an active ingredient.
 9. An agent forprotecting and promoting liver function comprising one of Brazilianlicorice extract and one or more constituents contained in the extractas an active ingredient
 10. The agent for protecting and promoting liverfunction as set forth in claim 9 wherein said one or more constituentscontain periandrins.
 11. Drugs comprising said agent for protecting andpromoting liver function as set forth in one of claims 8-10 as an agentfor protecting and promoting liver function active ingredient.
 12. Foodscomprising said agent for protecting and promoting liver function as setforth in one of claims 8-10 as an active ingredient for protecting andpromoting liver function.
 13. Food materials comprising said agent forprotecting and promoting liver function as set forth in one of claims8-10 as an active ingredient for protecting and promoting liverfunction.
 14. An anti-inflammatory agent comprising periandrins as anactive ingredient.
 15. An anti-inflammatory agent comprising one ofBrazilian licorice extract and one or more constituents contained in theextract as an active ingredient
 16. The anti-inflammatory agent as setforth in claim 15 wherein said one or more constituents containperiandrins.
 17. An immunosuppressant comprising periandrins as anactive ingredient.
 18. An immunosuppressant comprising one of Brazilianlicorice extract and one or more constituents contained in the extractas an active ingredient.
 19. The immunosuppressant as set forth in claim18 wherein said one or more constituents contain periandrins.
 20. Drugscomprising said anti-inflammatory agent as set forth in one of claims14-16 as an anti-inflammatory active ingredient.
 21. Cosmeticscomprising said anti-inflammatory agent as set forth in one of claims14-16 as an anti-inflammatory active ingredient.
 22. Foods comprisingsaid anti-inflammatory agent as set forth in one of claims 14-16 as ananti-inflammatory active ingredient.
 23. Food materials comprising saidanti-inflammatory agent as set forth in one of claims 14-16 as ananti-inflammatory active ingredient.
 24. Drugs comprising saidimmunosuppressant as set forth in one of claims 17-19 as animmunosuppressive active ingredient.
 25. Cosmetics comprising saidimmunosuppressant as set forth in one of claims 17-19 as animmunosuppressive active ingredient.
 26. Foods comprising saidimmunosuppressant as set forth in one of claims 17-19 as animmunosuppressive active ingredient.
 27. Food materials comprising saidimmunosuppressant as set forth in one of claims 17-19 as animmunosuppressive active ingredient.